Structural implications for the role of the N terminus in the ‘superactivation’ of collagenases

Abstract

For the collagenases PMNL‐CL and FIB‐CL, the presence of the N‐terminal Phe⁷⁹ correlates with an increase in proteolytic activity. We have determined the X‐ray crystal structure of the recombinant Phe⁷⁹‐Gly²⁴² catalytic domain of human neutrophil collagenase (PMNL‐CL, MMP‐8) using the recently solved model of the Met⁸⁰‐Gly²⁴² form for phasing and subsequently refined it to a final crystalographic R‐factor of 18.0% at 2.5 Å resolution. The PMNL‐CL catalytic domain is a spherical molecule with a flat active site cleft separating a smaller C‐terminal subdomain from a bigger N‐terminal domain, that harbours two zinc ions, namely a ‘structural’ and a ‘catalytic’ zinc, and two calcium ions. The N‐terminal segment prior to Pro⁸⁶, which is disordered in the Met⁸⁰‐Gly²⁴² form, packs against a concave hydrophobic surface made by the C‐terminal helix. The N‐terminal Phe⁷⁹ ammonium group makes a salt link with the side chain carboxylate group of the strictly conserved Asp²³². Stabilization of the catalytic site might be conferred via strong hydrogen bonds made by the adjacent, likewise strictly conserved Asp²³³ with the characteristic ‘Met‐turn’, which forms the base of the active site residues.